Methods to culture human pleural mesothelial (NHM) cells have been defined. Cultures are initiated by pelleting the mesothelial cells from pleural effusion fluid and inoculating the resuspended cells into dishes containing LHC basal nutrient medium supplemented with serum (3%), hydrocortisone (0.5 micromoles), insulin (5 micrograms/ml), epidermal growth factor (EGF) (5 ng/ml), transferrin (10 micrograms/ml), trace elements, and 2% chemically- reduced (factor-free) serum (FFS). Using this protocol, mesothelial cell cultures have been established from more than 200 donors. The cells can be subcultured at clonal density with a colony-forming efficiency of more than 10% and high density cultures can be subcultured four to six times before senescence. We have now established that many factors <EGF, transforming growth factor beta-1 (TGF-beta-1), TGF-beta-2, interleuken 1 (alpha) (IL- 1-alpha), IL-1-beta, transforming growth factor alpha (TGF-alpha), human platelet derived growth factor (hPDGF), porcine PDGF (pPDGF), fibroblast growth factor-acidic (FGF-a), FGF-basic (FGF-b), beta- interferon (Inf-beta), gamma-interferon (Inf-gamma), and cholera toxin beta will induce serum-starve cells to undergo one round of DNA synthesis in the absence of serum. However, for sustained growth, the medium must also be supplemented with high density lipids (HDL). Since human mesothelial cells have a peculiarly plastic cytoskeleton, we have characterized the affects of amosite fibers and code 100 glass fibers on the fidelity of division in these cells in an effort to understand the mechanism by which asbestiform fibers induce transformation and mesothelioma. Our results indicate that both amosite and code 100 glass fibers disturb the fidelity of cell division leading to the induction of aneuploid daughter cells. However, the mechanism of action of these two agents appears to be different. Amosite fibers induce chromosome clustering, suggesting an inhibition of mitotic tubulin formation and/or centriole separation. In contrast, code 100 glass fibers cause chromosome dislocation from the spindle at metaphase, suggesting an affect on centromere/kinetochore function and/or mitotic spindle function.